Biomimetic cell membrane‐coated poly(lactic‐ co ‐glycolic acid) nanoparticles for biomedical applications

Abstract Poly(lactic‐co‐glycolic acid) (PLGA) nanoparticles (NPs) are commonly used for drug delivery because of their favored biocompatibility and suitability for sustained and controlled drug release. To prolong NP circulation time, enable target‐specific drug delivery and overcome physiological barriers, NPs camouflaged in cell membranes have been developed and evaluated to improve drug delivery. Here, we discuss recent advances in cell membrane‐coated PLGA NPs, their preparation methods, and their application to cancer therapy, management of inflammation, treatment of cardiovascular disease and control of infection. We address the current challenges and highlight future research directions needed for effective use of cell membrane‐camouflaged NPs.

patients suffer from gastrointestinal (GI) symptoms, anemia, and loss of hair due to the effect of chemotherapy on normal cells, adding problems to patient's life. 3,4 Because of these serious side effects, the chemotherapeutic dose is kept low to reduce these complications; however, this may necessitate a prolonged course of therapy and even infective treatment with possible disease recurrence 5,6 and subsequent mortality. Targeted drug delivery has been proposed to address these limitations with the intent of directing drugs to specific tissue sites 7 ; this concept holds great promise for increased efficacy using lower doses with subsequent improved therapeutic outcomes and fewer side effects. 2,8 Recent advances in nanotechnology led to the development of nanoparticles (NPs) that can be decorated with targeting ligands, enzymes, polymers, and other biomolecules for directed delivery.
Nanoparticulate delivery systems comprise a paradigm shift, and engineered NPs can have specific physicochemical characteristics by altering surface charge, size, morphology, and surface hydrophilicity for enhanced accumulation in target tissues. 9,10 Despite significant advances in synthetic NP-based drug delivery systems, synthetic materials are foreign to the body, and can elicit nonspecific immune reactions. 11 Other challenges associated with these materials are their premature degradation and release of cargo, leading to rapid renal and hepatic clearance with insufficient drug being available for treatment, effective drug carriers must provide protection to their cargo by preventing premature degradation, prolonging in vivo retention, and protecting from immune surveillance, while preserving controlled drug release at the target site. 12 The use of biomimetic systems for drug delivery is an approach that may address some of these demands by imbuing cellular functions on synthetic particles. 13 By recapitulating the shape, movement and surface composition of cells, NPs may evade immune clearance to increase circulation time and protect the drug cargo, and if well-designed could also preserve the structure to improve drug release at the target site. Such agents would be more effective in vivo drug delivery tools. 14 Biomimetic systems employ cell membrane-camouflaged NPs that can be fabricated using top-down method, where core materials are coated with a cell membrane derived from natural cells. 15 Using this method, the resulting camouflaged NPs attain special functions including immune escape, 16 prolonged blood circulation, ligand recognition, and targeting, 17 which enable a wide range of applications of this system in drug delivery, 15 photothermal therapy, 18 vaccination, 19 and detoxifications. 9 Core materials used in the systems comprise a variety of materials that include polymeric NPs, silica NPs, gold nanocages, magnetic NPs, and liposomes. [20][21][22][23][24][25] Among these, polymeric materials can easily be prepared in various shapes having desired physical properties, modified with different functional groups suitable for various biomedical applications such as drug delivery and tissue engineering. 26 PLGA has been the most commonly used among polymeric NPs, because of its; (i) biocompatibility, (ii) approved status by both the United States Food and Drug Administration (FDA) and European Medicines Agency as a drug carrier, (iii) versatility for loading different types of drugs that may be water insoluble, and (iv) controllable biodegradation properties enabling tailored sustained release of drugs. 27 Because of recent successes in biocoating of NPs, there is increased interest that has led to more effective drug delivery tools, and we review these recent developments. A wide range of cell types have been used as sources of these coatings leading to different patterns in biodistribution. There have also been several methods for preparing the coated NPs that we discuss. The biomedical applications of these NPs are potentially far reaching, and we critically analyze and discuss the preparation and use of biomimetic NPs, highlight limitations of the technology and present future directions that would advance this strategy for effective drug delivery.

| Cell membranes
To achieve prolonged drug circulation, early research focused on the decoration of NPs' surface with synthetic polymers including polyethylene glycol (PEG), or encapsulating within liposomes or dendrimers to prevent their interaction with host environments. 28 For decades, PEGylation has been the method of choice for stealth coating. 29 However, recently it has been reported that PEGylated NPs are cleared from the circulation when second dose is administered because of an "accelerated blood clearance" phenomenon. 16 Because of the antibody production against PEG, scientists have been exploring new avenues to deceive the body's immune system. 11 Coating NPs with cell membranes is one of the most attractive approaches. This nature inspired strategy, leads to NPs that can efficiently interact with biological systems. 30 Redesigning the functions and structures of the cells used for coating NPs can lead to engineered surfaces with improved control of the biological processes. Imitating cellular processes can increase our understanding of molecular interactions, and by engineering the surface, we improve these interfaces. The synthetic engineered systems could provide a simple model to study the pathways underlying disease occurrence/development. These insights will lay a foundation for improved diagnosis and treatment. Development of biomimetic systems that imitate cellular processes constitute a tremendous research tool and a promising therapeutic approach. 31 In selecting cells as a source of lipid coating, consideration must be given to unique human proteins, antigens, that lack compatibility. The concepts used in transplantation biology including human-to-human transfer (allografts) and use of one's own cells (autograft) are relevant to coating NPs with cell membranes.
The cell membrane has a multitude of functions, one of which is protection from surrounding environment. The lipid bilayer that constitutes the major structure of the cell membrane contains a variety of proteins and carbohydrates that can be used as biomarkers, targeting ligands, or cloaking mechanisms. The molecules displayed on the cell surface impart specific functions and mechanisms of interaction with proximal cells and tissues. By wrapping synthetic NPs with the cell membrane, some of the functions are preserved and transferred to the NPs. 32 Cell membranes can be derived from different cell types, such as red blood cells (RBCs), 33 white blood cells (WBCs), 34 platelets, 35 stem cells, 36 and cancer cells, 37 and the characteristics of those cells can be transferred to the NP. The cell membrane disguises the particles, making them appear as "self" and thus not be rejected, or degraded, by the immune system. However, there are unique antigens on cell surfaces that can be immunogenic even if they are from human cells, for example, the Rhesus factor that is used in blood typing, and thus tissue matching remains relevant with these coated particles. 38 Nonetheless, the newly inherited features of the membrane-camouflaged NPs can be deployed for generating novel biointerfaces in the body that direct delivery as novel therapeutics. 39 In thinking about directed delivery, as it pertains to cells, particles and cell-particle hybrids, it is important to distinguish the mechanisms by which this can occur. The circulatory system distributes all materials in the blood around the body and the filtering organs such as liver and kidneys remove toxins, particulates and metabolic wastes and direct them toward excretion in the feces (via the liver) or the urine (via the kidneys). Circumventing the filtering process prolongs the circulation time and this is one objective for coating NPs. To direct an agent to a site of disease requires that the agent selectively accumulates at that site by sticking to or leaking into the target tissue. These mechanisms are possible with both particles and cells. Cells have the additional capability of sensing and actively migrating up a gradient, such as chemokine gradients; this active, energy-dependent mechanism is homing. Particles cannot actively home to a target tissue; they can selectively bind, accumulate, leak or otherwise be retained at the target tissue. It is the non energy-dependent characteristics of cell membranes such as the receptors they display, that may be transferred to NPs and optimizing these abilities will improve targeted delivery.

| RBC membrane-based strategy
RBCs have a circulation time of 120 days in the blood, and this long circulation time has attracted attention for development of drug carriers with prolonged circulation. The unique shape (flexible biconcave) of RBCs enables them to pass through tiny capillary networks. 40 The inherently non-immunogenic, biocompatible and biodegradable nature of RBCs has led to them becoming a lead candidate for intravascular delivery. RBCs can form natural compartments which provide prolonged protection to the encapsulated drug in blood stream, and can also sustain long release rates for small molecules. 41 A variety of proteins and glycans (carbohydrates) are displayed on the surface of RBC membranes that if transferred to NPs would help evade the immune system attack. 32 One such type of integrin-associated protein is cluster of differentiation (CD47) (often referred to as the "don't eat me" signal) that interfaces with its corresponding receptor on immune cells and enables RBC escape from destruction by circulating macrophages. 42 Iron oxide NPs were coated with RBC membrane to escape immune clearance through interactions with the signal regulatory protein-alpha (SIRP-α) receptor. Immune cells express SIRP-α which recognizes CD47 as a self-signal and prevent endocytosis of RBCs by defense cells. The RBC membrane maintained the CD47 glycoprotein after transfer to NPs, and coated NPs utilized the CD47-SIRP-α interaction for prolonged circulation. It was demonstrated that RBC membrane is a better alternative to the current gold standard PEG for prolonging the systematic circulation time of NPs. 16 However, RBCs lack active targeting capacity and related ligands, and thus additional functions must be added to direct delivery. Besides long circulating and selective targeting ability, controlled drug release is also required for an ideal drug carrier. In biomimetic NPs, the controlled release property can be conferred by the core materials. 24 2.1.2 | WBC membrane-based strategy Leukocytes or WBCs are the population of blood cells comprised of monocytes, lymphocytes and granulocytes. These cells are prevalent in lymphatic and vasculature and can extravasate into extravascular space by amoeboid movement. 18 Leukocytes are responsible for defense against infection and abnormal cells and are drivers of inflammation and antitumor immunity. During inflammation, selectins P and E are expressed on the surface of the endothelial tissue which facilitates binding of the leukocytes to the endothelium (blood vessel wall). The adherence of leukocytes to the endothelium can be used for targeting of drug carriers to sites of tissue damage or malignancy. 43 Many of the key biological activities of WBCs are mediated by glycoproteins on the outer surface of the WBC membranes including CD47, lymphocyte function-associated antigen-1 (LFA-1), and macrophage-1 antigen (MAC-1). Wrapping of NPs with WBC membranes transfers these cell surface markers to NPs, and some functions of the parent cells, such as sitespecificity and cellular self-recognition, are retained. 44 The surface proteins of WBCs imbue NPs with prolonged circulation, accumulation at sites of inflammation and disease, transendothelial migration and tumor tropism. 45 Multifunctional WBC membrane-based nanocarriers can be designed by employing the abilities native to leukocytes and they can be used for effective targeted delivery. These biomimetic NPs can be developed by either coating NPs with WBC membrane 46 or with leukosomes (specific extracted surface proteins) 47 or exosomes (leukocyte secreted extracellular vesicles) on core materials. 48 WBC membranes confer both camouflages to protect from immune destruction and some targeting functions, and an understanding of the cell surface proteins is important for effective NP delivery.

| Platelet membrane-based strategy
Platelets are small, nonnucleated cell fragments derived from megakaryocytes that circulate in the blood and are essential components for maintaining homeostasis. When blood vessels are injured, proteins such as collagen, which are present in the subendothelial layer, are exposed to platelets. After platelets bind to collagen, they release blood-clotting factors for the purpose to stop bleeding and promote wound healing. This property of platelets can be utilized to target vascular injury sites by coating NPs with platelet membranes. 49 Like   RBCs, different proteins are expressed on the surface of platelet   membranes such as CD47, CD55, and CD59, which can help to prevent the uptake of NPs by macrophages and can prevent unwanted immunological reactions. 50,51 The immune compatibility and prolonged blood residence time, owing to antigenic escape and both passive and active tumor target ability though binding to CD44 receptors upregulated on tumor surfaces, have been found to be useful when using NPs coated with platelet-derived membranes as cancer theranostics. 45 Platelet membrane-coated NPs can also be applied for targeted delivery and enzyme responsive release of plasminogen activators for fibrinolytic therapy and to minimize off-target systemic side-effects. Plasminogen activators convert plasminogen to plasmin, which act to break down the fibrin mesh in formed clot, and it is used in the treatment of occlusive vascular conditions such as stroke and heart attack. Platelet membrane-coated NPs were found to be capable of delivering their cargo to the clot within carotid artery in a thrombosis mouse model, without affecting the rest of the circulatory system. 52

| Cancer cell membrane-based strategy
Cancer cell membrane (CCM)-cloaked NPs represent an ideal candidate for oncological applications. Cancer cells are robust and are readily cultured for membrane extraction. Unlike other cell-derived membranes, CCMs may provide tumor-targeting ability which can be exploited in the development of novel therapies for primary tumors and their metastases. 53 The proliferation and metastases of cancer are primarily caused by advanced processes in which cancer cells escape the immune surveillance. CD47 molecule, which is overexpressed on CCM plays a key role in the process of immune eviction. CD47 molecule is particularly upregulated in some cancer cell lines such as MDA-231, 4T1 and MCF-7. Cell membranes rich in self-biomarkers and self-recognition proteins, can be isolated from cancer cells and wrapped around NPs, and enable them to evade the immune system in addition to providing tumor cell targeting. 32 Besides CD47, other proteins that help in cancer cell selfrecognition and tumor self-targeting are focal adhesion proteins, focal adhesion kinase, and RHO family proteins. 54 Cancer cells can also express neoantigens, proteins unique to the cancer cell, and these may invoke an immune response that would lead to elimination of cloaked NPs from the circulation, 55 57 The uniqueness of MSCs is attributable to the presence of surface molecules that are receptors for circulating ligands. These include receptors for cytokines, growth factors and chemokines, 58 as well as proteins for cellular interactions and cell-matrix adherence. 59 Signaling pathways activated by receptor-ligand interactions serve to connect the environment to cellular responses through expression of key metabolic and functional proteins. It has been reported that chemokines (CXCR1, CXCR2 CXCR4, CXCR5, CXCR6, CCR2, CCR7, CCR9, and CCR10) play a key role in homing, migration, and adhesion of MSCs to injured site or tumor. 60,61 In addition, MSCs possess ideal characteristics including non-immunogenicity, long systemic circulation, and inflammatory/ tumor-specific properties. 62 Certain stem cells such as MSC, hematopoietic stem cells, and neural stem cells can be exploited for drug delivery systems and carrying cargo to the tumor; these cells can secrete cytotoxic proteins that eliminate tumor cells. 11 As an auspicious source of cell membranes, MSCs can be extracted from different tissues and expanded in the laboratory. Furthermore, MSC-cloaked NPs can minimize blood clearance of circulating therapeutics by the reticuloendothelial system (RES) and increase cell specific uptake and enhance retention at tumor site. Although many of the functions present in MSCs do not transfer with the membranes, there are many passive interactions between the cell surface markers and the target tissues that are preserved. 63 The design of biomimetic drug delivery systems by surface modification of functional NPs with MSC membranes has advanced in many areas, 64 due to the unique properties of these cells.
Several cell types have been used as donors of cell membranes, and the key proteins that provide evasion and targeting properties are listed in Table 1.

| Poly(lactic-co-glycolic acid) NPs
Nanotechnology and polymer science have become the important areas of new developments in medical fields affecting a number of medical specialties and disease areas. 80 Biodegradable polymers are commonly used in both pharmaceutical and medical fields. 81 Among this group of polymers is PLGA, which is degraded mainly by hydrolysis, and traces of the breakdown products may be found in the urine. 82 PLGA has favorable biocompatibility properties, and has thus been used for a wide range of applications such as tissue regeneration, 83 osteofixation implants, 84 and drug delivery. 85 85,88 and physical properties that make is a well-suited carrier for drugs, proteins and other molecules including deoxyribonucleic acid (DNA) and ribonucleic acid (RNA). 89 The degradation time of PLGA can be altered from days to years by changing the molar ratio of lactide and glycolide, which is a favorable property of PLGA. Due to their safety profile and sustained release property, PLGA-based NPs are considered as effective nucleic acid delivery systems. 27,90 PLGA is prepared by the copolymerization of lactic acid and glycolic acid. The type and characteristics of PLGA are commonly specified by the molar ratio between these monomers.
PLGA polymers having lactic acid and glycolic acid in a ratio of 50:50 is the most common type of PLGA used for biomedical applications.
Glycolic acid monomer is less hydrophobic than the lactic acid mono- However, it can also decrease the encapsulation efficiencies of drugs.
Diblock polymers have shown improved release kinetics than PLGA alone. 89 The random polymerization of PLGA with other polymers will be beneficial; for example, combining PLGA with biodegradable photoluminescent polyester will make the system suitable for photoluminescence imaging. 97 The physical properties of PLGA nanostructure can be controlled by parameters specific to the preparation method employed. For example, concentration of PLGA used for preparation of NPs can determine the size of PLGA NPs to a certain extent. Surface decoration is another parameters that allows a certain control over particles' biodegradation, biocompatibility, blood half-life and, when applicable, targeting efficiency. 98 PLGA can be dissolved in wide range of organic solvent, depending on lactic acid and glycolic acid composition. If the ratio of lactic acid is high, the PLGA will dissolve by chlorinated solvents, such as chloroform or dichloromethane, and by water-miscible solvents, like tetrahydrofuran or acetone.
While, if ratio of glycolic acid is high, it will dissolve by fluorinated solvent, like hexafluoroisopropanol. 99 Finally, the glass transition temperature of PLGA can is above body temperature, between 40 and 60 C and this decrease with a decrease in the molecular weight. 100 Degradation of PLGA occurs in the following steps:   13 The field has been building on this pioneering work with erythrocyte membranes, by using other cell membranes to produce multifunctional membranecoated PLGA NPs.
Building on the theme, Hu et al. reported the use of plateletderived membrane-coated PLGA NPs. 51 In this study, platelets were selected because they have the distinctive property of sticking to surfaces of diseased or injured tissues. The resulting biomimetic NPs displayed platelet-imitating characteristics including selective binding to injured blood vessels of rodents and humans, increased binding to platelets with adherent pathogens. 51 An example of another cell type was neutrophil membrane-coated PLGA NPs. 111 With these NPs, it was possible to overcome the blood-pancreas barrier and achieve site-specific drug delivery. It has been postulated that homotypic cells (i.e., cells matching the disease) migrate to sites of the disease, and this was demonstrated using membranes from a homotypic HepG2 hepatocellular carcinoma (HCC) cell to direct chemotherapy to liver cancer (HCC). 112 Cell membranes may also facilitate uptake by resident cells in the diseased tissues as an additional mechanism for targeting. Human umbilical cord-derived MSC membrane-coated DOX containing PLGA NPs were fabricated using ultrasonication. 113 The in vitro uptake of membrane-cloaked NPs by cancer cells was threefold higher than that of bare PLGA NPs, leading to enhanced cancer cell cytotoxicity.
While cell membrane cloaking provides notable stealth-ability, additional selective targeting may further reduce their side effects and increase their therapeutic effect. The cell membrane-camouflaging approach has been progressively applied to more and more complicated biological systems using more varied and sophisticated engineering to incorporate a greater diversity of ligands and enhance their performance. The coated cell membrane can be directly modified using strategies such as covalent modification, non-covalent modification or enzyme-involved modification. 114 In covalent modification, amide bonds are formed between amine groups on the surface of the cell membrane and carboxylic acid groups present in the therapeutic moiety. 115 In non-covalent modification, lipid insertion is a commonly used, simple and stable approach. In this method, the functional group, related to lipids, can be spontaneously inserted into the phospholipid bilayer via hydrophobic interactions. If the functional moieties are linked with multiple hydrophobic interactions, then higher binding force is required. In contrary, the lipids insinuated into the exterior of cell membranes show good stability and strong molecular adherence to the target cells. 116 Along with lipid insertion, binding of certain peptides and antigens to the membrane surface proteins of cell membrane-coated NPs via ionic bond and hydrophobic interactions is another non-covalent modification strategy. However, this conjugation generally has a random distribution and the conjugated protein may lose function when changes to functional domains occur. 117,118 In enzyme-based modification, the therapeutic moiety is introduced onto the cell membrane via an enzymatic reaction, yielding high selectivity. For example, the enzyme, glycosyltransferase, can be used to introduce the therapeutic moiety CD44, derived from human MSCs, which selectively binds to P and E selectins on target cells. 119 Enzyme-involved reaction is also a kind of covalent modification. This approach has obtained a promising progress in cell membrane modification. But, it is difficult to utilize it to construct membrane-engineered cell membrane-coated NPs as each reaction demands the unique enzyme, which is challenging to be separated and purified from living cells. Also, it is hard to fully control the biochemistry reaction rate, which can be affected by different parameters, such as reaction temperature and membrane proteins. 120  These novel camouflaged nanocarrier combined the targeting ability of CAR-T cells with the advantages of nanoscale cores. 124

| Methods of preparation of PLGA-based cell membrane-camouflaged NPs
The fabrication of cell membrane-camouflaged NPs requires cell membrane extraction from the parent cells, fabrication of PLGA NPs, and coating of NPs using extracted cell membranes ( Figure 1). There are several published methods and approaches that are described below.

Extraction of cell membrane
Extraction of the plasma membrane from cells must be gentle to limit dissociation and denaturation of the membrane-associated proteins.
Cell membrane isolation typically includes a mild cell lysis followed by isolation of cell membranes, and varies according to cell type, specifically with respect to nucleated and nonnucleated cells.  should be optimized. 9 The sonication method can spontaneously produce core-shell NPs with reduced destruction of cell membrane structure. However, sonication is not appropriate for the production of cell membrane-coated NPs in large quantities. 133 Combined sonication and extrusion.  Table 3. The primary objectives of these modifications are to improve delivery to the target, increase circulation time for a more uniform delivery, and to reduce the systemic toxicity associated with conventional nanoparticulate drug delivery.

| Cancer therapy
Cancer treatment options include a wide variety of approaches and often include a combination of chemotherapy, surgery, radiotherapy, phototherapy, and immunotherapy with advances being made in combinatorial strategies as hybrid technologies. 137,138 Yet, there is no panacea and the limitations of conventional methods related to therapeutic performance, tumor targeting specificity and off-target toxicities are being addressed by modification of nanosized drug delivery systems.
Because these nanocarriers offer improved biosafety, performance, and bioavailability of a wide variety of therapeutic agents, they are defining the next generation of cancer therapies. 139 Biomimetic NPs with PLGA at their core can carry cytotoxic anticancer agents, or photoactivatable near-infrared dyes used in photodynamic therapy to the tumor, and are beginning to offer options to the oncologist for treating cancer patients. 140,141 Several of these are discussed below.   (Figure 3a). 146 The endocyto-  PLGA NPs were used for the delivery of verteporfin. 158 The use of platelet membrane wrapping helps to develop targeted solar irradiation photodynamic therapy and helps to limit the effect of reactive oxygen species in the area defined by the NPs. The platelet membrane envelope provides NPs with active targeting capability. Because of its inherent property to adhere to collagen type IV, binding of platelet membrane wrapped docetaxel loaded PLGA NPs to collagen type IV was evaluated.

| RBC membrane-coated NPs
Using fluorescent labeling, these NPs had significantly enhanced accumulation as compared to bare or RBC membrane-coated NPs, indicating the platelet membrane-type-specific adhesion to collagen IV.  Table 2.

| Inflammation
Inflammation is a protective response, activating the immune system to protect the body from hazardous effects of pathogens, foreign bodies or disease, 170 and the response may be acute or chronic. 171 Immune activation that persists beyond the initial insult, which is due to a self-antigen, or a result of nonspecific stimulation of immune cells leads to chronic inflammation that becomes the pathology. Such inflammatory conditions are characteristic of several disease conditions such as rheumatoid arthritis (RA), 172 GI inflammatory disorders, 173 cancer, 174 pneumonia, 175 and CVDs. 176 (Figures 5f,g).
Besides, survival time of the mice was extended compared to the control group (Figure 5h). 180 Granulocyte macrophage colony stimulating factor distinguishes monocytes into premature dendritic cells which then travel toward inflamed tissue and process tumor antigens to both recruit and activate T-lymphocytes. 181 Many of these processes require adherence molecules on the immune cells for them to be retained at the tumor location, therefore, cell membrane-cloaked NPs may provide an opportunity for the targeted delivery of anticancer therapeutics. 182 The vasculature of tumors is disrupted, irregular and leaky, thus cells and particles accumulate at the tumor site due to the effect known as EPR. Once cells and particles leak into the tumor microenvironment,  162 Arsenic trioxide 1. Enhanced circulation time and sustained release of arsenic trioxide and reduced toxicity. 2. RBCs-PLGA arsenic trioxide NPs showed lower cytotoxicity than arsenic trioxide solution to normal 293t kidney cell lines and an antitumor effect against HL60 cells by CCK8 assay. 163 2 WBCs Doxorubicin 1. Enhanced cellular uptake and cytotoxicity of doxorubicin loaded nano ghosts as compared to non-coated NPs in breast cancer (MCF-7) cell lines (in vitro). 164 3 Cancer cells DiD fluorophore 1. In cancer immunotherapy, the facilitation of multiple antigens combined with immunological adjuvants was due to uptake of membrane bound tumor antigens for downstream immune activation. 2. In anticancer therapy, cancer cell membrane coating provides homotypic binding mechanism.

| GI inflammatory conditions
The inflammatory response in the GI tract is used to eliminate pathogens and toxins and is usually self-limiting. 191   pancreas and selectively distributed in the pancreatic tissues better than noncoated NPs. Pancreatic acinar cells produce phospholipase A2 (PLA2), which is known as trigger of acute pancreatitis. Therefore, macrophage membrane-coated PLGA NPs doped with melittin (a short peptide that can trap PLA2 to attack the membrane) and MJ-33 (PLA2-specific inhibitor) were developed to direct therapy to PLA2 expressing cells in the treatment of acute pancreatitis (Figure 7a,b). 195 The MJ-33 and melittin work as PLA2 obstructor and PLA2 attractant respectively. These NPs could counteract PLA2 activity in a dosedependent manner, and depress PLA2-activated inflammatory responses (Figure 7c-f). Compared to control groups, notable reduction in edema score, counts of necrotic acinar cells and hemorrhagic area in the pancreatic tissue were observed with NPs (Figures 7g-i).

| Cardiovascular diseases
CVD has the highest morbidity and mortality worldwide, 196 and disorders such as cardiac and cerebral ischemia 197 and atherosclerosis 198 are largely associated with vascular inflammation. 199 The foremost causes of disability and death in the developed world have long been CVDs and their sequelae, 200 and prevention has been the focus of many treatment strategies. Although the cholesterol-lowering drugs have been used successfully for the past 25 years, novel treatment protocols focusing on vessel wall inflammation have also been examined, [201][202][203] and targeted therapy can be employed. 8,[204][205][206] 3.

| Atherosclerosis
Atherosclerosis is a gradual inflammatory disease that has a lipid and fibrous artery wall build-up, and is a significant cause of mortality and morbidity. 207 Currently, atherosclerosis is treated with both medicines and surgical intervention. Treatment of early-stage atherosclerosis is restricted to general oral medications. 208 Even though procedures such as stenting are effective for treating progressive atherosclerosis, these procedures are associated with side effects, such as restenosis and thrombosis. 209,210 On the other hand, nanotechnology enables the specific targeted delivery of therapeutic compounds, has the potential to increase efficacy and safety, [211][212][213] and NPs with a biomimetic natural surface can avoid the RES and can be used for specific targeting. 214,215 Atherosclerosis is a disease driven largely by macrophages. When these cells accumulate in the vessel wall, they secrete cytokines that recruit additional immune cells and metalloproteinases leading to thrombosis. 216 Conventional therapies have been ineffective due to poor delivery and the need for long-term administration at high doses that are associated with serious side effects. 217   Platelets have an inherent affinity for atherosclerotic plaques, and RAPA loaded PLGA NPs coated with platelet membranes demonstrated targeted drug delivery and significantly reduced the progression of atherosclerosis in mouse models. 77 Lumbrokinase is another drug used to treat thrombotic diseases, but nonspecific delivery is associated with hemorrhagic (bleeding) complications. 221 Therefore, platelet membrane-coated PLGA NPs loaded with lumbrokinase were developed to risk and overcome the short half-life of the drug. These NPs demonstrated selective adherence to thrombotic plaques, had thrombolytic activity, and reduced adverse effects compared to free drug.
In another study, macrophage membrane coating was applied to the surface of RAPA-loaded PLGA NPs. 222

| Ischemia
Stroke is the third leading cause of mortality in the United States, causing almost 150,000 deaths per year. 226 Ischemic stroke is the most frequent type of stroke and happens when blood supply to the brain is interrupted due to thrombosis. 227 The presence of biological barrier (BBB) is the major hurdle in clinical management of stroke.
In later stages of stroke, the integrity of BBB is partially disrupted, but it remains largely intact within the therapeutic window and may not allow the transfer of pharmacologically required drugs for effective treatment. Nanotechnology can enhance the brain penetrability, but existing approaches have been shown to be inadequate. 228 Therefore, biomimetic NPs may provide means to cross BBB, and increase the accumulation of drugs at the sites of disease, as a result they can be used as innovative diagnostic systems for early stage stroke diagnosis, for example to detect several biomarkers (ROS and neurotransmitters). 78,229,230 Ischemia of the heart is another leading cause of death worldwide, and for the diagnosis of myocardial ischemia reperfusion injury, platelet membrane-coated PLGA NPs have been developed. 231 Coated NPs showed enhanced in vitro and in vivo performance as compared to noncoated NPs. Furthermore, localization to the target area was achieved as indicated by echo signal intensity, which was significantly higher in the risk area as compared to remote areas of the myocardium in rat models. An immunofluorescence assay and ex vivo fluorescence imaging was used to validate these findings.

| Infectious diseases
Infectious diseases are accountable for majority of the hospitalization and are a prime cause of mortality and morbidity around the globe. 237 The effective treatment of infectious disease remains a challenge due (MRSA) are a significant concern. 247 S. aureus secretes α-toxin, a membrane destructive pore-making toxin, which has various cellular targets such as leukocytes, epithelium, platelets, and endothelium. 248 Helicobacter pylori infects over half of the global population, 249 and is a major cause of gastritis, digestive ulcers, and gastric cancer.
Currently, a triple therapy comprised of clarithromycin, proton pump inhibitor and an antibiotic either metronidazole or amoxicillin is the suggested treatment protocol. 250 Drug resistance mutations in H. pylori have been found which make it resistant to these antibiotics, 251 new and effective treatments are being sought. 252 These include gastric epithelial cell membrane-coated clarithromycin containing PLGA NPs, which were found to exhibit superior binding to H. pylori and accumulation at the target site as compared to noncoated NPs. 193  For cell membrane extraction from various cells, differential centrifugation and density gradient centrifugation methods are generally used. 278 However, the centrifugation parameter and choice of cell fragmentation methods have not been clearly established yet. The stability of cell membrane without nutrient supply and cytoplasmic support is still an issue. The risk of bacterial contamination is another issue that may complicate platelet membrane, which are usually stored at 22-24 C for 5-7 days. 278 Yield of coating of NPs with cell membranes is another challenge with cell membrane production, for example, for high yield of WBC, platelet and stem cell membranes are difficult to achieve. It has been estimated that to coat 1 mg of PLGA NPs, up to 3 Â 10 9 of platelets are needed. In this regard, a simplified method with high yield of cell membrane is urgently needed. 279 Large-scale manufacturing of cell membrane-camouflaged NPs is a major limitation for wider use of coated NPs. Cell membrane coating technology is primarily dependent of membrane extrusion and sonication. 12 Both these methods have pros and cons. For example, sonication is characterized by high production efficiency but low uniformity, while membrane extrusion has low production efficiency and high uniformity. Cell factories, fully automated cell culture sys- In addition to cell membranes, there are also challenges related to PLGA drug carriers, despite their use already over 30 years. These are related to formulation-related aspects pertaining to drug loading and release. One of the key issues related to fabrication of biosimilar PLGA-based drug products is the complexity of the manufacturing process. Slight changes in the manufacturing process, involving quality control/assurance (QC) systems, can greatly influence the bioactivity, efficacy, safety, and stability of the product. Polymer degradation and drug release are greatly influenced by the physicochemical properties of PLGA such as molecular weight, particle size, porosity, drug polymer interaction, and glass transition temperature. 281 The higher entrapment efficiency of water soluble drugs in hydrophobic PLGA matrix is the major limitation. Grafting PLGA with block polymers having amphiphilic sites will greatly improve the encapsulation efficiency of water soluble drugs. 281  which shows the superior efficacy of extrusion process than sonication in the formation of a full cell membrane coating. The integrity of cell membrane coating affects the endocytic entry mechanism of coated NPs. NPs with a high degree of coating (>50%) display individualized cellular internalization, whereas those with low degree of coating (<50%) require aggregation before internalization. 284 Partially coated NPs were found to enter cancer cells via a cooperation mechanism based on appropriate aggregation of NPs. 284 Therefore, careful evaluation of the ratio of full cell membrane coating will enhance tumor targetability.
The asymmetric biological characteristics of cellular membranes, differences between the cytoplasmic and outer surfaces, means that the right-side-out orientation of cell membrane coating is also a vital for effective therapies. As seen in the case of RBC membrane coating on PLGA NPs, a strong negative charge exists on the extracellular side of RBC membrane than on the intracellular cytoplasmic side. 64 Therefore, electrostatic interaction is favored between intracellular surface of RBC membrane and negatively charged PLGA NPs, which biased the right-side-out orientation. This mechanism will not benefit positively charged NPs.
For the development of versatile cell membrane NPs, certain modifications to the plasma membrane are inevitable and may induce undesirable adverse effects. For example, excessive use of immune cell membrane coating may interact with immune system, leading to release of pathological mediators and may induce inflammation. 12 In case of platelet membrane coating, the pro-inflammatory effect has been reported to promote the development of atherosclerosis. 285 Biocompatibility is another concern of cell membrane-coated NPs.
Although short-term biocompatibility has been demonstrated in different studies, 56 still the cell membrane extraction should be undertaken with great care to avoid potential risks. Limitations related to manufacturing of cell membrane-coated PLGA NPs can be overcome with the use of 3D printing, microfluidics and particle replication in non-wetting templates (PRINT). Recently, PRINT technology has enabled precise manufacturing of engineered particles with uniform biodegradable material cores and independent control over their chemical composition, shape and size to impart desired therapeutic benefits to the product. 197 Although many applications have been explored, there certain issues that need better understanding. For example, the mechanism of drug release from cell membrane-coated NPs is still unclear. 178 Moreover, in comparison to synthetic materials, the purity and quality of coated NPs are hard to control, and their safety is also uncertain especially for CCM coated NPs, which can induce a harmful immune response. 286   Abbreviations: AD, Alzheimer disease; BBB, blood-brain barrier; DSS, dextran sulfate sodium; NPs, nanoparticles; PLGA, poly(lactic-co-glycolic acid); RBC, red blood cell; VEGF, vascular endothelial growth factor. leaky vasculature. Second, co-loading/administration of vasodilators or constrictors with nanovesicles to modulate tumor blood flow will enhance the EPR effect. Third, the anchoring of ECM-degrading enzymes, like hyaluronidase, collagenase, and so forth to membrane bilayer will augment the abnormal tumor ECM and can assist diffusion and spatial distribution of nanovesicles in the interstitial space. 287 Regarding PLGA nanoparticles can entrap both water soluble and insoluble drugs, protect the drugs from degradation, provide sustained release and can increase therapeutic efficacy by improving the pharmacokinetics and pharmacodynamics of the active pharmaceutical ingredients. Therefore, merging of natural cell membranes and synthetic PLGA NPs core is an exciting area of investigation that may lead to improved delivery of therapies for the treatment of pathological conditions such as cancer, inflammation, CVDs and microbial infections.
Cells have much to teach us about targeted delivery and using their membranes on synthetic nanoparticles will lead to discovery of innovative approaches and new opportunities in medicine.

CONFLICT OF INTEREST
The authors declare no conflict of interest.

DATA AVAILABILITY STATEMENT
The data that support the findings of this study are available from the corresponding author upon reasonable request.